(611e) Buffer Matters: Investigating the Influence of Buffer Selection on Protein Retention in Ultrafiltration Processes

Ultrafiltration and diafiltration (UFDF) are integral processes in the formulation of therapeutic proteins, crucial for achieving final concentration and buffer exchange. However, maintaining high product yield can pose a significant challenge for some products. Yield losses can occur due to sieving losses, membrane adsorption, hold-up volume, and product quality degradation. While efforts have been made to address some of these challenges through the development of hydrophilic membranes and strategies to enhance product recovery, the impact of buffer selection on protein retention remains underexplored. This study investigates the influence of different buffers on protein retention during ultrafiltration, using bovine serum albumin (BSA) as a model protein. Despite previous research emphasizing the role of pH, ionic strength, and ion binding on BSA retention, the specific effects of buffer species have yet to be quantified. Buffer species are known to affect protein conformational stability, enzymatic activity, and thermal stability, which in turn can impact ultrafiltration behavior. Dynamic light scattering (DLS), differential scanning calorimetry (DSC), and circular dichroism (CD) are employed to elucidate conformational changes that may influence protein retention. BSA retention was significantly reduced in phosphate buffer compared to that in acetate, citrate, and histidine due to conformational changes that cause a small reduction in the effective hydrodynamic diameter of the protein. BSA retention was greater at pH 6.5 than at 4.8 due to a combination of conformational changes and electrostatic repulsive interactions. This work provides important and new understanding of the factors controlling protein retention, which is essential for mitigating sieving losses in UFDF processes and optimizing biotherapeutic formulation strategies.